Method and apparatus for performing intra-operative angiography

ABSTRACT

Method for assessing the patency of a patient&#39;s blood vessel, advantageously during or after treatment of that vessel by an invasive procedure, comprising administering a fluorescent dye to the patient; obtaining at least one angiographic image of the vessel portion; and evaluating the at least one angiographic image to assess the patency of the vessel portion. Other related methods are contemplated, including methods for assessing perfusion in selected body tissue, methods for evaluating the potential of vessels for use in creation of AV fistulas, methods for determining the diameter of a vessel, and methods for locating a vessel located below the surface of a tissue.

CROSS-REFERENCE TO RELATED PATENT APPLICATIONS

This application is a continuation application of U.S. application Ser.No. 11/106,154, entitled METHOD AND APPARATUS FOR PERFORMINGINTRA-OPERATIVE ANGIOGRAPHY and filed on Apr. 14, 2005, which is acontinuation application of U.S. application Ser. No. 09/744,034,entitled “METHOD AND APPARATUS FOR PERFORMING INTRA-OPERATIVEANGIOGRAPHY” which issued as U.S. Pat. No. 6,915,154, which is anational stage entry of PCT application PCT/US00/22088, filed on Aug.11, 2000, which claims priority to provisional application number60/155,652 which was filed on Sep. 24, 1999.

TECHNICAL FIELD OF THE INVENTION

This invention generally pertains to procedures for observing blood flowthrough the cardiovascular system of an animal.

BACKGROUND OF THE INVENTION

Disease and injury affecting the cardiovascular system in animals, andparticularly humans, are commonplace in today's society. One suchdisease is atherosclerosis. This disease is characterized by partialblockage (stenosis) of a blood vessel, typically by a narrowing of oneor more arteries. In its most severe form, the vessel narrows to thepoint that it becomes completely blocked (occluded). In coronaryarteries, stenosis and occlusion often manifest themselves in the formof severe chest pains and, potentially, myocardial infarction (heartattack). Not limited to coronary arteries, atherosclerosis can alsoaffect the peripheral vasculature, i.e., arteries (and veins) thatcirculate blood throughout the arms and legs, the carotid arteries,i.e., arteries that carry blood to the brain, and intracranial arteries,i.e., arteries that distribute blood within the brain.

One therapy commonly employed in an effort to overcome the effects ofatherosclerosis in coronary and peripheral vessels is bypass graftsurgery. During this procedure, a vascular graft, e.g., a vein or arteryor, alternatively, a flexible artificial tube, is surgically inserted ina manner that permits blood to bypass the stenotic or occluded portionof a native vessel. Perhaps the best-known example of bypass graftsurgery is coronary artery bypass graft (CABG) surgery. In CABG, agraft, commonly a saphenous vein or internal mammary artery, isharvested or dissected from the patient, respectively, and then locatedwithin the patient to permit blood flow to bypass the stenotic oroccluded vessel portion. Alternatively, or in addition thereto, a graftmay be used to permit blood to flow directly from the aorta to alocation downstream of a stenotic or occluded portion of an artery.

The success of bypass grafts, at least in terms of clinical improvement,depends in significant part upon the ability of the treated vessel toremain free of occlusions over both the short- and long-term. Thisfreedom from occlusions is commonly referred to as vessel patency. Poorpatency in the first few months after surgery is thought to be theresult of various factors, with the following believed to be the mostsignificant: poor blood circulation, poor coronary arterial runoff,injury to the graft during preparation or faulty surgical technique.

While cardiac surgery in recent years has focused on strategies tominimize trauma to the myocardium, these strategies may increase thelikelihood of problems if used during vessel grafting procedures. Forexample, while surgical techniques now permit CABG to be performed on abeating heart to minimize trauma, there exists a concern relating to thequality of the resulting graft. The use of limited access incisionsduring CABG procedures has been developed for, at least, therevascularization of the left anterior descending artery using a leftinternal mammary artery, with the hope of faster recovery, a shorterhospital stay and reduction in cost. However, this method has alsoraised concerns relating to graft quality. Indeed, there exist reportsof early failure in grafts completed using limited access incisions.

Other issues affecting CABG procedures are diagnostic in nature, andinclude relatively slow and inaccurate identification of stenotic andoccluded vessels during the initial phase of CABG procedures (as some ofthese vessels lie within the heart tissue which inhibits visualidentification), and an inability to quickly and accurately determinethe extent of blood flow through the relatively smaller downstreamvessels (and, more generally, whether the graft was successful inrestoring blood flow to affected tissue) after the graft is completed.

Arterial patency issues may arise in therapies that do not includegrafts. For example, patency evaluation is desirable in carotid arteriesduring and after an endarterectomy, in cranial vessels during and afterneurosurgery, and in the context of kidney hemodialysis, wherein anassessment of AV fistula patency is desirable. While vessel patencyinformation in these contexts may be obtained using X-ray technology,the disadvantages mentioned previously remain.

The extent of blood flow within a particular tissue or portion thereof,commonly referred to as perfusion, is important in connection with thediagnosis and treatment of a variety of ailments. For example, aperfusion analysis would be desirable in the context of a treatmentdesigned to reduce undesired blood flow into tissue, e.g., halting bloodflow into a tumor. At present, MRI may be used to obtain perfusioninformation, but this information is imprecise and only available aftertreatment is completed. This lessens the probability that a physicianwill be able to identify and remedy problems during that same procedure,thereby precluding the need for a subsequent remedial procedure.

Another affliction that requires treatment of the circulatory system isrenal failure. In many cases of renal failure, it is desirable to createan AV fistula to provide vascular access for hemodialysis. The fistulais created by joining an artery and vein by a surgical procedure,providing a vessel having a relatively high rate of blood flow. WhileX-ray technology can be used to assist the physician in determiningwhether the creation of a properly functioning fistula is possible, andthe type of fistula that should be created, the technology suffers fromthe previously mentioned limitations.

In view of the foregoing, a need exists for a diagnostic procedure thatpermits a physician to evaluate the patency of a particular vessel, andparticularly vessels that have undergone an invasive procedure such as abypass graft procedure. A further need exists for a method of quicklyand accurately locating a particular stenotic or occluded vessel, suchas a coronary artery during the initial phase of CABG surgery. Inaddition, improved methods for evaluating the extent of blood flowdownstream of a graft are needed, e.g., in coronary arteries andperipheral vasculature, as are more accurate methods for determining theextent of blood perfusion in selected body tissue. A need also existsfor an improved means of identifying candidate vessels for AV fistulas,and of obtaining information relevant to a determination of the type offistula that should be created in a patient with renal impairment.

BRIEF SUMMARY OF THE INVENTION

The present invention meets the forgoing and other needs by providing,in one aspect, a method for assessing the patency of an animal's bloodvessel, advantageously during an invasive procedure in which the vesselis treated. The method comprises the steps of administering afluorescent dye to the animal; obtaining at least one angiographic imageof the vessel portion; and evaluating the at least one angiographicimage to assess the patency of the vessel portion.

A related aspect provides for assessing blood flow in a portion oftissue in an animal wherein the tissue is a candidate for an invasiveprocedure, is undergoing an invasive procedure, or has undergone such aprocedure, comprising identifying the tissue portion in the animal;administering a fluorescent dye to the animal; obtaining at least oneangiographic image of blood flowing through the tissue portion; andexamining the at least one angiographic image to assess blood flow inthe tissue portion.

A further aspect of the present invention permits a physician toaccurately determine the extent to which a selected portion of bodytissue, e.g., heart tissue, tumor, is well perfused, to assist in theidentification and diagnosis of improperly (or properly) perfusedtissue. The method comprises the steps of selecting a portion of bodytissue to be analyzed; administering a fluorescent dye to the patient;obtaining at least one angiographic image of the selected tissue; andexamining the at least one angiographic image to assess the extent ofblood flow within the selected portion of body tissue.

In a related aspect, the present invention provides a method forevaluating chemical agents and other proposed therapies in terms oftheir effect on vasculature. The method comprises obtaining a firstangiographic image of selected vasculature; administering a therapeuticagent; obtaining a second angiographic image of the selected vasculatureon a subsequent day; and comparing the first and second angiographicimages to determine if there is any change in vascular density over thattime period.

In another aspect of the present invention, a method of locating, in ananimal, at least one vessel (or portion thereof) residing beneath thesurface of vascularized tissue is provided. The method comprises thesteps of administering a fluorescent dye to the animal; obtaining atleast one angiographic image of the vasculature located beneath thesurface of the tissue; and examining the at least one angiographic imageto locate the at least one vessel residing beneath the surface of thetissue.

In a further aspect, the present invention provides an apparatus fordetermining the diameter of a blood vessel. More specifically, theapparatus comprises: a device that emits radiation capable of causingfluorescent dye to fluoresce; a camera capable of capturing theradiation emitted by the fluorescing dye within the blood vessel as anangiographic image comprised of a plurality of pixels; and a computercomprising a software program that calculates the diameter of a bloodvessel by comparing the number of pixels that correspond to the bloodvessel diameter with the number of pixels associated with a preselectedunit of measurement.

These and other features and advantages of the present invention willbecome apparent upon review of the following figure and detaileddescription of the preferred embodiments of the present invention.

BRIEF DESCRIPTION OF THE DRAWING

FIG. 1 illustrates in schematic form a preferred embodiment of theapparatus of the present invention.

DETAILED DESCRIPTION OF THE INVENTION

The methods of the present invention are claimed and described herein asa series of treatment steps. It should be understood that these methodsand associated steps may be performed in any logical order. Moreover,the methods may be performed alone, or in conjunction with otherdiagnostic procedures and treatments administered before, during orafter such methods and steps set forth therein without departing fromthe scope and spirit of the present invention. Further, it iscontemplated that the term animals as used herein includes, but is notlimited to, humans.

Turning now to one aspect of the present invention, a method is providedfor analyzing the patency of a portion of an animal's blood vessel. Themethod comprises the steps of administering a fluorescent dye to theanimal; obtaining at least one angiographic image of the vessel portion;and evaluating the at least one angiographic image to assess the patencyof the vessel portion.

Illustrative of the vessels whose patency may be evaluated in accordancewith the inventive method include coronary arteries, the peripheralvasculature, carotid arteries, intracranial vessels and AV fistulas. Anevaluation of vessel patency may be conducted qualitatively by a visualinspection of the images and, if desired, quantitatively by obtaining ameasurement of vessel diameter, wherein a substantially uniform diameterof a particular vessel portion's lumen is desirable.

Advantageously, vessel patency may be determined during an invasiveprocedure. For purposes of this and other aspects of the presentinvention, an invasive procedure is one in which one or more incisionsare made in the tissue of an animal, or entry of an instrument into anorifice of an animal is undertaken, to diagnose or treat an afflictionor condition that directly or indirectly affects vasculature or tissue.The invasive procedure should be understood to continue until theincisions are sutured, or the instrument is withdrawn from the animal,respectively.

By way of example, this aspect of the invention contemplates aphysician, during a single invasive procedure, obtaining angiographicimages of a coronary artery both prior to and after treatment (e.g.,bypass). In this way, the physician is able to quickly evaluate thepatency of the treated vessel. This is beneficial because it allows aphysician, upon noting a problem in the treated vessel, to take remedialmeasures during the same invasive procedure, sparing the animal from thetrauma associated with a subsequent remedial invasive procedure.

Examples of vessel portions that may benefit from use of the inventivemethod include, but are not limited to, vessels that have been subjectedto: repair (due to injury, aneurysm and/or malformation) or bypass (ofcoronary arteries or peripheral vasculature); endarterectomies;intracranial surgery; creation of AV fistulas; and surgical proceduresconducted using an endoscope or related devices.

Illustrative of the types of repair include, but are not limited to:lacerated vessels closed by suture or adhesive; removal of an aneurysmor other vessel malformation by removing the undesired portion of avessel followed by either joining the two remaining ends of the vesselto one another, or the interposition and subsequent joining of a naturalor synthetic vessel graft to the remaining vessel ends.

Bypass is commonly used when a portion of a blood vessel, typically astenotic or occluded portion, requires circumvention. Bypass includes,but is not limited to, attaching the ends of a graft vessel at locationsupstream and downstream of the stenosis, occlusion or other problem, aswell as grafting one end of a relatively healthy artery onto theundesired vessel at a location downstream of the stenosis, occlusion, orother problem. One specific example of the latter is a procedure whereinone end of a healthy artery from the chest wall is grafted onto acoronary artery downstream of a stenotic or occluded portion thereof.The inventive method is preferably utilized in surgery involving thebypass of coronary arteries, e.g., CABG surgery.

When bypass is undertaken, an anastomosis, i.e., the junction of thenative and graft vessels, is created. The patency of anastomoses is ofparticular interest to physicians. In a preferred aspect, the inventivemethod contemplates the assessment of the patency of anastomoses, morepreferably during the invasive procedure, and most preferably while theheart remains beating.

A further aspect of the present invention provides a method forassessing blood flow in a portion of animal tissue wherein the tissue isa candidate for an invasive procedure, is being or has undergone aninvasive procedure. In the latter case, an evaluation of the extent ofblood flow through vasculature located downstream of a treated vesselassists a physician in assessing the success of the treatment. Themethod comprises identifying a portion of animal tissue; administering afluorescent dye to the animal; obtaining at least one angiographic imageof blood flowing through the tissue portion; and evaluating the at leastone angiographic image to assess blood flow in the tissue portion.

This method may advantageously be used in the assessment of flow incoronary arteries and peripheral vasculature, and is preferably usedduring an invasive procedure. In one preferred aspect, the methodcontemplates obtaining an angiographic image of vasculature locateddownstream of a particular blood vessel, e.g., a coronary artery, thathas undergone treatment, e.g., bypass, to assess the success of thebypass procedure. In another preferred aspect, the method contemplatesobtaining an angiographic image of vasculature located downstream of aparticular peripheral vessel that has undergone treatment, e.g.,peripheral vessel bypass, wherein the image is obtained without incisingthe skin overlaying the downstream vasculature. In the latter aspect,the treated peripheral vessel and/or downstream vasculature ispreferably located at a depth below the skin surface that permits thevasculature of interest to be assessed. Preferably, this depth is atleast about 0.5 cm, and more preferably at least about 1 cm, below theskin surface.

This aspect of the present invention further contemplates assessing theblood flow in other body tissues including, but not limited to, muscle,stomach, liver, intestine, bladder, esophagus, lung, kidney and braintissue. Angiographic images may be obtained beneath the surface of thesetissues to a depth not exceeding that which permits the vasculature ofinterest to be evaluated. Again, and preferably, this depth is at leastabout 0.5 cm from the surface of any of the foregoing tissue, and morepreferably at least about 1 cm, with access to the tissue by endoscopebeing a preferred route. This method may be used in connection with avariety of invasive procedures, such as those that assess whetherinternal bleeding has been halted. For example, a physician will be ableto readily determine whether a surgical treatment successfully haltedbleeding in what was previously a bleeding ulcer.

The inventive method further provides a means of evaluating varioustherapies, wherein the success of such therapies is indicated at leastin part by the extent of blood flow in or about a particular tissue. Themethod contemplates obtaining a first angiographic image of a selectedtissue; administering the therapy (e.g., a proposed therapeuticcompound) to the animal; obtaining a second angiographic image of thesame selected tissue at a later time (e.g., hours, days or monthsthereafter); and comparing first and second images to determine whetherthere is any change in vascular density and/or blood flow within thetissue. One use of this method is in the evaluation of angiogenic andanti-angiogenic agents, as well as in the research of such potentialtherapies. For example, an endoscope may be used to evaluate the impact,if any, of a particular therapy on decreasing the flow of blood intoand/or through tumors, such as lung or colon tumors.

In another aspect of the present invention, a method of locating a bloodvessel residing below the surface of vascularized tissue, e.g., astenotic or occluded artery or vessels suitable for the creation of anAV fistula, is provided. The method comprises the steps of administeringa fluorescent dye to an animal; obtaining at least one angiographicimage of the vasculature located beneath the surface of the tissue; andexamining the at least one angiographic image to locate at least onevessel residing beneath the surface of the tissue.

As the method permits ready visualization of vessels located down to atleast about 0.5 cm, and preferably down to at least about 1 cm below thetissue surface, a physician is potentially able to complete a bypass orother coronary procedure involving the location of stenotic or occludedvessels residing below the tissue surface in less time, simply due tothe time saved in locating the vessel to be treated.

In the context of renal failure, the method provides a means of locatingarteries and veins that are suitable for the creation of an AV fistula,as well as providing information that assists a physician in determiningwhich type of fistula to create based upon the structure of thevasculature. In a preferred aspect, the method permits angiographicimages of peripheral vasculature located down to thepreviously-described depths to be obtained without requiring an incisioninto the skin to expose the vasculature of interest.

Angiographic images obtained in the absence of an incision may also beuseful in assessing a peripheral (upper and lower extremities)vasculature bypass (by evaluating the blood flow through the vasculaturedownstream of the bypass), and in assessing endothelial dysfunctionthrough the nail bed (by assessing the extent of blow flow throughcapillaries located under the nail bed).

The angiographic images obtained in accordance with the various aspectsof the present invention depict the lumen (space) inside the arteriesand veins located within the subject tissue. A relatively thick lineindicates a major artery, whereas a relatively thin line indicates asmaller artery. A line of substantially uniform thickness indicates avessel that is free of atherosclerotic plaques. In contrast, a line thatis ragged, or that becomes thinner in certain sections, indicates thepresence of stenosis, while a discontinuation of a line indicates thepresence of an occlusion.

In yet another aspect, the present invention provides an apparatus andrelated method of providing images of high resolution that permit aphysician to determine vessel diameters down to about 30 μm and less.This aspect of the invention will be discussed in more detail insubsequent paragraphs.

In order to obtain an image in accordance with the various aspects ofthe present invention, a fluorescent imaging agent is administered tothe patient. The fluorescent agent should be selected so that when itpasses through the vasculature of interest, at least one useful image ofthe vasculature can be obtained. Fluorescent dyes emit radiation of aknown wavelength when excited by radiation of a particular wavelength.The radiation emitted by the excited dyes is detectable, and may becaptured by a suitable device that converts the radiation into aviewable image.

While any fluorescent dye may be used that provides an image asdescribed herein, indocyanine green (ICG) (IC-GREEN™, CARDIO-GREEN™,marketed by Akorn, Inc.), analogue members of the tricarbocyanine dyes,and mixtures thereof, are preferred. ICG is preferred because it isreadily available, and has long been approved for administration tohumans for ophthalmic angiography, cardiac output analysis and otherindications.

The wavelengths for both absorption and emission radiation associatedwith such dyes are well known, and will not be repeated herein. By wayof example, however, as the peak absorption and emission of ICG lies inthe range of 800-850 nm, a radiation source emitting such wavelengthsshould be used to obtain one or more images of the vessels or tissue ofinterest.

Typically, the fluorescent agent is administered in a composition thatincludes a pharmaceutically acceptable carrier. The composition shouldbe administered in an amount, and the fluorescent agent present at aconcentration, sufficient to provide the degree of detail desired in theimages. Advantageously, the agent is present in an amount of from about1 to about 10 mg/ml, preferably from about 3 to about 7 mg/ml, and morepreferably about 5 mg/ml of the composition, with the carrierconstituting the balance thereof.

The carrier, which advantageously solvates but which may merely emulsifyor suspend the agent, is provided to enhance the administration of theagent to a patient. Administration is typically accomplished viaparenteral, IV injection, or other suitable means, with IV injection ofthe composition as a bolus being preferred, with the carrier beingselected in view of the desired mode of administration.

Illustrative carriers that may be used include water, saline, alcohols,glycerin, polyethylene glycol, propylene glycol, polysorbate 80, Tweens,liposomes, amino acids, lecithin, dodecyl sulfate, lauryl sulfate,phospholipid, Cremophor, desoxycholate, soybean oil, vegetable oil,safflower oil, sesame oil, peanut oil, cottonseed oil, sorbitol, acacia,aluminum monstearate, polyoxyethylated fatty acids, povidone andmixtures thereof. Advantageously, the carrier comprises water and/orsaline.

Optional components that may be present with the agent in thecomposition include tonicity and/or pH adjusters, e.g., NaOH, HCl,phosphate buffers, Tris buffer and the like.

The composition that comprises the agent may initially be provided inany suitable formulation, for example, as a lyophilizate forreconstitution before use, or as a liquid pre-mix, in a vial or syringe.

After administration of the imaging agent, a device capable of excitingany of the agent that may be present in the vasculature or tissue ofinterest, and a device capable of detecting the radiation emitted fromany such agent, are activated. While each device may be provided in aseparate housing, they may also be combined in a single housing withoutdetracting from the present invention. Turning to FIG. 1, the device forexciting the agent advantageously comprises a laser 1 which emitsradiation at a wavelength, that causes any of the agent located withinthe vasculature or tissue of interest 3 irradiated thereby to emitradiation of a particular wavelength.

Lasers that are capable of providing radiation suitable to excite theagent sufficiently to permit detection of emissions are well known tothose skilled in the art (e.g., Magnum 3000, Lasiris St-Laurent, Québec,Canada), and as such will not be described in detail herein. Generally,however, the devices comprise a laser driver and diode, andadvantageously a bandpass filter 5. The filter 5 assists in optimizingimage quality by ensuring that the radiation reaching the vessel is of asubstantially uniform wavelength, i.e., the wavelength that causes theagent to fluoresce.

As the field of illumination provided by the laser alone is insufficientto radiate an anastomosis or other relatively large area, the laseradvantageously includes optics 7 which diverge the laser light to coverthe area of interest. By way of example, it has been found that opticsthat provide for even irradiation of a 7.5 cm×7.5 cm area will besufficient to irradiate most anastomoses. Such optics are well known,and will therefore not be described in detail herein. Preferably, theoptics should permit variation in the field of illumination, as it issometimes desirable to concentrate the laser radiation on a relativelysmall area to enhance image resolution.

In a further optional enhancement, the laser output may be pulsed,synchronized with the camera image acquisition rate by use of a devicesuch as a pulse generator 18. This reduces the amount of laser radiationreceived by the vessel or tissue while retaining image quality.

Devices capable of detecting emissions from imaging agents, andparticularly the preferred fluorescent dyes, are also well known.Advantageously, a camera capable of obtaining multiple images over aperiod of time, such as a CCD camera 2 (e.g., Hitachi KP-M2, KP-M3), maybe used to capture the emissions from the imaging agent. The cameraselected, of course, should be one capable of capturing radiation of thewavelength emitted by the imaging agent. Preferably, the camera shouldcapture such images at a rate of at least 15 images/sec, and morepreferably at a rate of at least about 30 images/sec. The camera mayalso be fitted with a bandpass filter 6 to prevent capture of radiationother than that emitted by the imaging agent.

The camera focus may be by automatic or manual means. Further, and ifdesired, the camera may include a lens system 8 that enables an area ofinterest to be magnified. Preferably, the use of such a lens system isswitched to the laser so that, when the lens system is engaged, thefield of illumination provided by the laser is correspondingly reducedto match the field of view provided by the lens system. The result ofthis coordination is enhanced resolution. Polarizing filters 14 a, 14 bmay also, if desired, be fitted to the laser and/or camera to enhanceresolution.

Advantageously, a distance sensor 9 (e.g., WTA 24, SickOptic-Electronic, Inc., Eden Prairie, Minn.) is included as part of theapparatus. This sensor, which preferably incorporates a visual display 9a, provides feedback to a physician so that the laser and camera can belocated a distance from the vessel or tissue of interest that is optimalfor the capture of high quality images, thereby minimizing the need forfocusing of the camera during the procedure.

The relative positioning of the camera and laser can also affect imageclarity, also referred to as visual noise. Preferably, and as shown inFIG. 1, the laser is located at an angle Θ of less than about 85°, andmore preferably between about 20° and 70°, with respect to the axes ofthe laser and camera. Introducing the laser radiation into the bodycavity at these angles reduces the amount of glare entering the cameraarising from the liquid present in the cavity.

While the camera and laser may be located external to the patient, asshown in FIG. 1, it is also contemplated that at least one endoscope maybe used to obtain images of the type described herein. For example, inthis aspect of the invention, the endoscope would be inserted into thebody, through an incision and/or body cavity, and positioned adjacentthe area of interest. A first instrument, typically a laser optic fiber,would be inserted into the endoscope, and used to provide radiation atan appropriate wavelength to cause any of a previously administeredimaging agent within the subject vessel or tissue to emit detectableradiation. A second instrument inserted into the endoscope that wouldpermit an image of the radiation-emitting agent within the vessel ortissue to be obtained. For example, an optical device connected to a CCDcamera, such as those used to perform a colonoscopy, may be readilyadapted for use in conjunction with the endoscopic procedurecontemplated by the present invention. The manufacture of a suitabledevice in view of the disclosure provided herein is believed to bewithin the skill of the ordinary artisan, and will not be described indetail herein.

Preferably, the camera relays the captured images to ananalog-to-digital converter 10 (typically a card located within PC 15),and then through image-capture and processing software running on a PC15. A digital image of the fluorescing agent (which corresponds to thelumen of the vein, artery and/or anastomosis of interest) may then bedisplayed on a monitor 11, and recorded by the PC or a peripheral devicein any suitable medium, e.g., hard drive, optical disc, magnetic tape,or the like. The camera may also direct images directly to a television12/VCR 13 system, wherein the images may be displayed in real timeand/or recorded for playback at a later time. Preferably, the monitorand/or television are located in the surgical suite, permittingreal-time viewing of various aspects of the treated and surroundingvessels. A printer 16 may also be connected to the camera, PC and/or VCRto permit a hard copy of one or more angiographic images to be obtained.

Analog-to-digital converters are well known. These devices, as theirname implies, convert the series of analog images captured by the camerato digital images. Image processing software is also well known, with avariety of software presently available that is capable of analyzing thetreated and adjacent vessels.

In practice, it is preferred that the camera, laser and video monitor belocated opposite the surgeon, to ensure that the surgeon has maximumspace to position the device relative to the patient. The remainingcomponents may be placed in any convenient location. Preferably, thelaser, camera and/or video monitors are mounted on one or more armaturesthat provide freedom of movement along the x, y and z axes to providemaximum maneuverability, and which remain in a desired position afterplacement.

In a preferred aspect, the image-capture and processing software is ableto provide a measurement of the diameter of a blood vessel, e.g., thediameter of the treated portion of a vessel and the end portions of theoriginal vessel adjacent the treated portion. While a number ofdifferent methodologies may be used to provide this measurement, onesuch method follows. As the invention contemplates that the camera bepositioned in a different location for each patient, or to obtain imagesof more than one vessel in a single patient, the software advantageouslyincludes a calibration algorithm that permits an operator to assign adistance to a specified number of image pixels. While calibration can becompleted using any suitable method, one method involves the use of acapillary tube of a known inner diameter filled with a fluorescent dye,e.g., ICG. The dye in the capillary tube is excited by radiation from alaser, and the resulting image of the fluorescing liquid detected by thecamera, and processed by the software, is used to assign a length to thenumber of pixels that correspond to the inner diameter of the capillarytube.

The software preferably includes a further feature that selects theoptimal images for analysis. The need to have such a feature is basedupon the relatively fast flow of the imaging agent through the tissue ortreated vessel of interest under normal conditions. Because the timingof the passage of imaging agent (if any is able to pass therethrough)through the tissue or vessel of interest cannot be precisely determined,there exist a number of leading and trailing images acquired before andafter the images of interest. The software is preferably capable ofdetermining the relative contrast of one image with another, and in thismanner selects those frames with the greatest contrast for analysis,i.e., in the case wherein the agent is able to enter the vessel ortissue of interest, those frames in which the imaging agent is presenttherein and emitting detectable radiation. This selected series ofimages may then be analyzed to determine the diameter of the treated (orany other vessel) at a particular location, as well as the rate andvolume of blood flow through the treated vessel and adjacent originalvessel.

Software may also be used to compare images of pre- and post-treatmentvessels to determine the relative flow rate of blood at or downstream ofthe treatment site. This comparison is accomplished by calculating andcomparing the area of fluorescence (i.e., number of pixels associatedwith the fluorescing dye) in pre- and post-treatment images associatedwith a preselected section of the vessel, and/or comparing the relativeaverage maximum fluorescent intensity of a preselected section of thevessel in each such image. A greater number of pixels, or greateraverage maximum fluorescent intensity, respectively, in thepost-treatment images indicates improved blood flow in the preselectedvessel section as a result of the treatment.

Similarly, the invention permits the diameter of a vessel to becalculated and compared both before and after stimulation, e.g., theadministration of acetylcholine. This comparison is significant, becausean increase in vessel diameter demonstrates that the vessel hasmaintained endothelial function, which is a positive indication offuture vessel patency.

The advantages of the present invention are further illustrated by thefollowing example. The particular details set forth therein should notbe construed as a limitation on the claims of the present invention.

EXAMPLE

This example demonstrates the use of a preferred apparatus of thepresent invention in observing the flow of a fluorescent dye through aparticular vessel, i.e., a mouse femoral artery, and langendorffperfused heart, and also demonstrates the ability of the apparatus todetermine the diameter of a mouse femoral vessel under both normalconditions and under the influence of topically applied acetylcholine.

In this example, a fluorescent dye (ICG) was injected into the vascularbed (via jugular cannulation in the mouse: via an infusion line in thelangendorff perfused heart) and excited using radiation from a lasersource (806 nm). The fluorescence (radiation) emitted by the dye (830nm) was captured as a series of angiograms using a CCD camera. Thecamera relayed the angiograms to analog-to-digital conversion softwarerunning on a PC that digitized the angiograms. The digitized images werethen analyzed both qualitatively (by viewing the monitor) andquantitatively. One example of quantitative evaluation that wasundertaken was the determination of the mouse femoral artery diameterusing software comprising a sub-pixel edge detection system running onthe PC.

The foregoing fluorescence imaging technique was used on the mousefemoral artery in vivo. A more detailed explanation of each component ofthe apparatus, preparation of the animal, injection of ICG, andanalytical method, are set forth in the following paragraphs.

The laser device included an SDL-820 Laser Diode Driver (SDL Inc., SanJose, Calif.) that maintained a continuous wave output with an averagecurrent of 3.95 A, and an SDL-2382-P1 laser diode (SDL Inc.). The laserdiode was used to illuminate the area of interest and excite the ICGdye, thereby inducing fluorescence in the region being imaged. A laserdiode was used because, unlike an incandescent light source, a laseremits photons in a narrow frequency range, and thus eliminates the needfor an excitation filter and the associated problem of heat dissipation.Because the laser-emitted wavelengths are limited, the excitation filtercan be eliminated, improving the fluorescence. Consequently, a higherproportion of the light emitted from the laser diode is of thewavelength absorbed by ICG. It was found that use of an 800DF20 bandpassfilter (Omega Optical Inc., Brattleboro, Vt.) in conjunction with thelaser light source improved the results by selectively passing photonsemitted at 806 nm (i.e., the wavelength at which ICG is excited).

The angiographic images were collected using a KP-160 video camera(Hitachi Denshi, Ltd., Tokyo, Japan). The KP-160 camera was selectedbecause it is highly sensitive in the near-infrared region of theelectromagnetic spectrum (which is also where ICG fluoresces), thusoptimizing the capture of radiation emitted from the excited ICG. An845DF25 bandpass filter (Omega Optical Inc., Brattleboro, Vt.) wascoupled to the camera to exclude all photons that were not of thewavelength associated with ICG fluorescence. The laser diode waspositioned at a 45° angle to the area of investigation in order tominimize specular reflectance (i.e., glare) arising from surface waterfrom entering the camera. Glare is a major source of visual noise duringimaging.

An analog-to-digital converter (752×480 pixel, 8-bit image processor,Model PIXCI-V4, EPIX Inc., Buffalo Grove, Ill.) was employed to digitizethe composite video signal output from the camera.

After each IV injection of an ICG dye bolus, a series of 264 interlacedimages was collected at a rate of 30 per second.

The mouse was prepared by inducing anesthesia in an induction box usingisoflurane (Ohmeda Pharmaceutical Products, Mississauga, ON, Canada) (4%in medical air, 4 L/min) and maintained by use of a facemask providingisoflurane at a rate of 1.5-2.0% in medical air (400 mL/min). During theexperiment, the mouse was positioned on a thermostatted water blanket,with body temperature being monitored by a rectal temperature probe. Tofacilitate imaging of the vessels of interest, the thoracic, abdominaland inguinal areas of the mouse were shaved, the mouse positioned on itsback, and the skin over the femoral vasculature was resected to exposethe vasculature of interest. The jugular vein was cannulated using apiece of stretched PE10 tubing filled with saline containing 50 Uheparin/mL.

After the mouse was prepared, a 10 μl bolus IV injection of ICG wasadministered, followed by an IV injection of 50 μl of saline solution.To prepare the bolus, 4 μg/ml of clinical grade ICG (CARDIO-GREEN™) wasdissolved in sterile aqueous solvent within one hour of injection. Allinjections were administered via the cannula established in the jugularvein. The saline was used to flush the line and to ensure passage of anintact bolus through the femoral vasculature, producing a sharpwavefront.

Image analysis was performed using XCAP for Windows 95/98/NT version 1.0(EPIX Inc., Buffalo Grove, Ill.). The image processing algorithmincluded the following steps.

1. Selection of vessels of interest. The anatomy of the vasculaturevaries between animals. Consequently, it was necessary to developcriteria for the selection of an area of interest. This process beganwith the positioning of the camera. The camera was positioned so thatthe field of view included the femoral artery and its branches. For thepurposes of image analysis, the vessels of interest were the femoralartery and the branches that provided the highest resolution and thegreatest degree of branching, usually tertiary or quaternary.

2. Calibration. The positioning of the camera with respect to the areabeing imaged varied with each animal, and it was therefore necessary tocalibrate the camera for every image collected. A small diameter (320μm) capillary tube (TSP320450; Polymicro Technologies, LLC, Phoenix,Ariz.) filled with ICG was used to calibrate the images. The imageprocessing software includes a built-in calibration function that allowsthe specification of a set of pixel co-ordinates and the assignment of auser-defined value to the distance between these co-ordinates. Thesoftware's edge detector was used to determine the co-ordinates of theedges of the dye fluorescing in the capillary tube. The inner diameterof the capillary tube, in microns, was then assigned to the “length” ofthe distance between these points. Because this is a built-in feature ofthe software, all subsequent measurements in all frames of the imagewere stated in microns, rather than pixel units.

To avoid distortions due to camera movement or other stochasticphenomena, every image was calibrated. The advantages of this techniqueare that the same method was used to measure the calibration device aswas used to measure the vessel, and the calibration device is measuredin the same frame under the same optical conditions as the vessels.

3. Measurement of diameter using sub-pixel edger. All vessel diameterswere measured using the built-in sub-pixel edger.

4. Selection of frames based on edge strength. Analysis of ICG imagesentails the selection of frames for analysis. The need to select framesis a consequence of the fast rate of ICG flow through the femoral arterywith respect to the rate of image acquisition. This results in a leadingand trailing sequence of frames that were acquired before and after ICGwas detectable in the area being imaged. Edge strength, which isautomatically calculated by the edge detector in our software, is ameasure of the relative strength of the edge, i.e., the ratio of thevalue of the pixels on one side of the edge to the value of those on theother side. The ratio is highest when the contrast is greatest, whichcorresponds to the greatest intensity of ICG fluorescence. The vesselsthat were measured have two edges, thus ten frames in which the productof the edge strengths was the greatest were selected for analysis.

After the foregoing was completed, the vessel diameters and standarderrors were calculated as described above. Student's t-test for pairedvalues was applied to determine the statistical significance between themeasurements (border of significance, p=0.01.)

Preliminary data on the effects of different size vessels in the mouse(femoral artery) are given in the Table. The data confirms the abilityto monitor changes in small vessels (e.g., 58 microns) when even a lowconcentration of acetylcholine (0.01 μM) is applied.

TABLE Effects of Acetylcholine Acetyl- choline concentra- VesselDiameter (microns) tion control 0.01 μM .01 μM 1.0 μM 10.0 μM Primary92.7 ± 1.2 58.2 ± 1.3 61.5 ± 1.7 58.3 ± 1.5 64.6 ± 1.5 Secondary 69.4 ±0.3 67.0 ± 1.3 75.1 ± 1.2 90.0 ± 1.8 75.0 ± 1.4 Tertiary 57.5 ± 0.7 42.9± 0.6 44.9 ± 0.6 47.1 ± 1.2 42.9 ± 0.8 p < 0.05

The foregoing demonstrates the ability of the present invention toobserve the flow of blood through a vessel, to determine the diameter ofa vessel, and to monitor changes in the reactivity of a vessel after theadministration of acetylcholine.

All of the references cited herein, including patents, patentapplications, and publications, are hereby incorporated in theirentireties by reference. Further, and unless otherwise indicated,references to a single component, structure or step herein should beconstrued as also including more than one such component, structure orstep, i.e., at least one or one or more.

While this invention has been described with an emphasis upon preferredembodiments, it will be obvious to those of ordinary skill in the artthat variations of the preferred embodiments may be used and that it isintended that the invention may be practiced otherwise than asspecifically described herein. Accordingly, this invention includes allmodifications encompassed within the spirit and scope of the inventionas defined by the following claims.

1. A device for monitoring movement of a fluorescent dye contained inblood moving through a cardiovascular bypass graft and vasculatureconnected to the graft of a subject during a surgical procedure, thedevice comprising: an illumination source emitting radiation of which aportion is absorbable by the moving fluorescent dye; a camera having animage capturing device configured to capture images of fluorescentradiation emitted from the fluorescent dye moving through the bypassgraft and connected vasculature while the subject's heart is beating, atan acquisition rate of at least 15 angiographic images per second, thecaptured images including at least an optimal image of fluorescent dyeinitially moving through the bypass graft and connected vasculature, aswell as images leading and trailing the optimal image; wherein thefluorescent dye has a peak absorption in the range of 800 to 850 nm; anda display device that transforms the captured images into viewableimages.
 2. The device according to claim 1, wherein the camera capturesimages at an increased acquisition rate of at least 30 images per secondand the dye is indocyanine green.
 3. The device according to claim 1,wherein the illumination source and the camera are positioned withrespect to one another such that an angle between an optical axis of thecamera and an axis of a radiant energy beam produced by the illuminationsource is less than 85°.
 4. The device according to claim 1, wherein theillumination source is a laser, and the device further comprises opticspositioned to diverge a radiant energy beam produced by the laser. 5.The device according to claim 4, wherein the laser and the camera arepositioned with respect to one another such that an angle between anoptical axis of the camera and an axis of the radiant energy beam fromthe laser is between 20° and 70°.
 6. The device according to claim 4,wherein the optics are adjustable, permitting variation in a field ofillumination.
 7. The device according to claim 4, further comprising abandpass filter positioned relative to the radiant energy beam to limitthe radiant energy beam to one of a substantially uniform wavelength. 8.The device according to claim 4, wherein the laser is pulsed andsynchronized with the acquisition rate of the camera.
 9. The deviceaccording to claim 1, further comprising a bandpass filter positioned toprevent the camera from capturing radiation other than that emitted bythe fluorescent dye.
 10. The device according to claim 1, wherein thecamera comprises a lens system for magnifying a field of view.
 11. Thedevice according to claim 10, wherein the lens system is capable ofbeing engaged by the laser to correspondingly adjust a field ofillumination provided by the laser as a function of the field of view ofthe camera.
 12. The device according to claim 4, further comprising adistance sensor that provides distance information from at least one ofthe camera and the laser to the cardiovascular bypass graft.
 13. Thedevice according to claim 4, wherein the optics engage the laserresulting in irradiation of a 7.5 cm×7.5 cm area.
 14. A method forevaluating movement through a cardiovascular bypass graft andvasculature connected to the graft of a fluorescent dye contained inblood moving through the bypass graft and vasculature of a subjectduring a surgical procedure, comprising: injecting the fluorescent dyehaving a peak absorption and emission in the range of 800 to 850 nm intothe blood of the subject; illuminating with an illumination source thecardiovascular bypass graft and vasculature connected to the graft withradiation that will be absorbed by the fluorescent dye; capturingradiation emitted from the fluorescent dye within the cardiovascularbypass graft and vasculature connected to the graft as an angiographicimage with a camera that captures images at an acquisition rate of atleast 15 images per second; and evaluating the captured images to assessthe extent of blood flow in the bypass graft and the vasculatureconnected to the graft.
 15. The method according to claim 14, whereinthe camera captures images at an increased acquisition rate of at least30 images per second and the dye is indocyanine green.
 16. The methodaccording to claim 14, wherein the fluorescent dye is excited with aradiant energy beam from the illumination source, and the illuminationsource and the camera are positioned with respect to one another suchthat an angle between an optical axis of the camera and the radiantenergy beam is less than 85°.
 17. The method according to claim 14,wherein the illumination source is a laser that emits a radiant energybeam, and the laser is associated with optics positioned to diverge theradiant energy beam to cover the cardiovascular bypass graft and thevasculature connected to the bypass graft.
 18. The method according toclaim 15, wherein the fluorescent dye is excited with a laser that emitsa radiant energy beam, and the laser and camera are positioned withrespect to one another such that an angle between an optical axis of thecamera and the radiant energy beam is between 20° and 70°.
 19. Themethod according to claim 17, wherein the optics are adjustable,permitting variation in a field of illumination of the illuminationsource.
 20. The method according to claim 17, wherein a bandpass filteris positioned relative to the radiant energy beam to limit the radiantenergy beam to one of a substantially uniform wavelength.
 21. The methodaccording to claim 17, wherein the laser is pulsed and synchronized withan acquisition rate of the camera.
 22. The method according to claim 17,wherein a bandpass filter is positioned to prevent the camera fromcapturing radiation other than that emitted by the fluorescent dye. 23.The method according to claim 17, wherein the camera comprises a lenssystem for magnifying a field of view.
 24. The method according to claim23, wherein the lens system is capable of being switched to the laser tocorrespondingly adjust a field of illumination provided by the laser asa function of the field of view.
 25. The method according to claim 17,wherein the optics illuminate an illumination area of a 7.5 cm×7.5 cm.